Development of the nervous system PDF

19/01/2024 Development of the nervous system Where opportunity creates success 1 Learning objectives Reviewing the concept of Gastrulation Describe neural development Explain the formation of the neural plate and t...

19/01/2024 Development of the nervous system Where opportunity creates success 1 Learning objectives Reviewing the concept of Gastrulation Describe neural development Explain the formation of the neural plate and tube Understand the patterning of the nervous system Describe the process in synapse formation 2 1  19/01/2024 Why is neurodevelopment important? Developmental Origins of Health and Disease (DOHaD) 3 Ninja Nerd https://www.youtube.com/watch?v=BtLyik7oAxc 4 2  19/01/2024 Contents ❖ Neural development ❖Early brain development Research methods in neural development Differentiation of the forebrain Development of ectoderm Differentiation of the midbrain Primary neurulation Differentiation of the hindbrain Secondary neurulation Differentiation of the spinal cord ❖ Neural crest How are boundaries set up? ❖Construction of neural circuits Failure of NCC Neuronal polarization ❖Histogenesis within the CNS Synapse formation Pattering of the neural tube ❖ Neurovascular diseases Cell lineage of CNS Neuronal migration in the CNS 5 Review of gastrulation Gastrulation→ begins with the primitive streak Primitive streak→ site where the cells in the epiblast layer start to migrate Formation of three distinct layers: the 3 germ layers: 1. Ectoderm: skin and nervous system 2. Mesoderm: bones and muscle 3. Endoderm: internal organs 6 3  19/01/2024 Gastrulation movements Gastrulation in the chick. Fluorescently labelled cells along the primitive streak allow you to see the movement of cells during gastrulation. - The red cells are more anterior and you can see them moving laterally and anteriorly. - The green cells, which are more posterior, move in a more lateral direction. 7 Formation of the notochord Notochord→ cylinder of mesodermal cells Functions: 1. It is positioned centrally in the embryo with respect to both the dorsal-ventral (DV) and left- right (LR) axes 2. Send signals to the ectoderm causing cells to differentiate (thicken the ectoderm→ neuroectodermal precursor cells) 8 4  19/01/2024 Neural development From the first 2 weeks to stem cell regeneration of adult neural tissue In between this there is the generation of complex structure The nervous system develops: - from the anterior to the posterior - later from the inside towards the outside The nervous tissue develops in conjunction with other tissues. 9 10 5  19/01/2024 Research methods to study neural development Microdissection and transplantation Checking gene transcription and expression: – Add protein soaked beads – RT-PCR, PCR – Add cells expressing protein – In Situ Hybridisation: whole Overexpression: introducing plasmids by mount and section, fluorescent and antibody electroporation and microinjection, viruses – Microarray Controlled expression: cre-lox Analysing Protein expression: Knock down: RNAi – Immunohistochemistry and immunocytochemistry Knock out: CRISPR electroporation, – Western Blotting plasmids Histology In vitro and in vivo imaging: cell culture and tracking of gfp expression, brainbow 11 Development of ectoderm Neurulation begins at ~18d, shortly after gastrulation Neurulation: process of folding the neural plate into a neural tube. This tube forms the basis for the brain, spinal cord and associated neurons 1. Primary neurulation→ ectoderm is divided into three sets of cells: - neural tube, which will form the brain and spinal cord, - the externally positioned epidermis of the skin - the neural crest cells 2. Secondary neurulation→ the tube forms by hollowing out of the interior of a solid precursor 12 6  19/01/2024 Primary neurulation 1. Formation of the neural plate Neural plate→ midline ectoderms which contains the thicker cells Border of the neural plate (neural crest)→ BMP Ectodermal→ subdivided into 2 developmental lineages: neural and non-neural 2. Shaping the neural plate Narrower and longer Cells forming the neural plate migrate toward the midline and also become longer along the anteroposterior axis and narrower laterally 13 3. Lateral folding of the neural plate Elevation of each side of the neural plate along a midline neural groove Formation of hinge points - Dorsolateral hinge points - Medial hinge points 4. Neural tube closure Cells from the 2 folds merge (fusion of the 2 sides) – Extension of lamellipodia and filopodia – Cells interlock and fuse Neural crest begin to separate from the neural tube Detachment from the ectodermal layer 14 7  19/01/2024 Begins around 21- to 22-day-old embryo Zip-like closure at 3 sites in mouse, 5 in human - Caudally - Cranially: two additional discontinuous sites of closure Neuropores→ unclosed cephalic and caudal parts of the neural tube 15 Consequences of NTC Failure NTC 2nd most common defect worldwide Spina bifida, anencephaly Varies in severity, defects can be significantly reduced with supplementation of Folic Acid EARLY in pregnancy Can be detected by increased levels of α-fetoprotein in maternal serum and the amniotic fluid What does Folate do? 1. Role in DNA, RNA, protein and lipid methylation- affects activity and functionality 2. Involved in DNA repair and replication via production of DNA precursors. It is an enzyme co-factor 16 8  19/01/2024 Cadherin change to detach the neural tube Changes allow the edges of the tube to join together and to separate from the ectoderm. E-cadherin→ N-cadherin N-CAM→ provide for fine-tuning of intercellular connections 17 EphA7 and Ephrin A5 bind to maintain closure of the neural tube Ephrins and Eph receptors are transmembrane proteins: activating signalling pathways (protein- tyrosine kinases) Receptors (EphA7) and ligands (Ephrin A5) expressed along the neural folds Reliant on cell-cell contact to signal and work Knockout of EphA7 and Ephrin A5 can lead to failure of NT closure. Other ligands may be responsible for regional closures. 18 9  19/01/2024 Secondary neurulation Caudal to posterior neuropore (evolved in vertebrates with longer tails?/ no prominent in humans) Formation of a rod-like condensation of mesenchymal cells Occurs via cell migration and condensation Mesenchymal cells change and become epidermal in identity. Attach to neural tube and cavity becomes continuous 19 Instructions Join at vevox.app Or search Vevox in the app store ID: 128-025-112 Where opportunity creates success 20 10  19/01/2024 Join: vevox.app ID: 128-025-112 POLL OPEN Which of the following techniques could be used to investigate RNA expression in a human blood sample? 1. Cre-lox 2. Immunohistochemistry 3. RNAi 4. RT-PCR 5. Western blot 21 Join: vevox.app ID: 128-025-112 POLL OPEN Presence of alpha fetoprotein in the amniotic fluid indicates a deficit in which process? 1. Angiogenesis 2. Gastrulation 3. Interneuron migration 4. Neural tube closure 5. Synaptic pruning 22 11  19/01/2024 2 minute break 23 Neural Crest From the prosencephalon to the future sacral region Nonneural lineage from the ectoderm Cells located along the lateral margins of the neural plate Migrate to other positions around the body 1. Trunk crest subdivision migrate along the ventral pathways between the neural tube and the dermomyotome - Pathway 1→ sympathoadrenal lineage - Pathway 2→ dorsal root ganglia - Pathway 3→ pigment cells 24 12  19/01/2024 2. Cranial crest subdivision Migrate along the dorsolateral pathway between the somites and the ectoderm Neural crest cells migrate from the mesencephalon into the head and from rhombomeres: - r1 and r2→ first pharyngeal arch - r4→ second arch - r6 and r7→third arch 3. Circumpharyngeal Neural Crest Transition between cranial and trunk neural crest 4. Cardiac crest Separate the outflow tract of the heart into aortic and pulmonary segments 25 Neural Crest Cell migration Images of neural crest cells in chick embryos moving away from the neural tube and into pharyngeal pouches. 26 13  19/01/2024 Failure of NCC migration Intestinal Aganglionosis (Hirschsprung's Disease or Megacolon) Genetic disease Nerves are missing in some areas of the digestive system Surgery removing affected areas Neurofibromatosis Genetic disease Tumors grows in the nervous system 3 types Surgical removal of the tumors Neuroblastoma Genetic mutation 100 children/year in UK Adrenal gland tumours or along the spinal cord. 27 28 14  19/01/2024 Histogenesis within the Central Nervous System 29 Proliferation Within the Neural Tube Neuroepithelial cells - high degree of mitotic activity - within the neural tube DNA synthesis→ external limiting membrane Orientation of the mitotic spindle predicts the fate of the daughter cells: - Metaphase plate perpendicular to inner surface of neural tube→ cell migrate in tandem back toward outer side - Metaphase plate parallel to inner surface of neural tube - daughter cell that is closer to the inner surface migrates away and remains a proliferative progenitor cell - other daughter cell progresses to the next stage in the neural lineage 30 15  19/01/2024 Patterning of the neural tube Neural patterning→ biological process by which cells in the developing nervous system acquire distinct identities according to their specific spatial positions Controlled by signalling gradients along the axis of the developing nervous system: - dorsal–ventral - rostral-caudal 31 Dorsal-Ventral axis Compartments of neural progenitor cells that lead to distinct classes of neurons Roofplate: Dorsal signal→ bone morphogenetic proteins (BMPs) Commissural neurons, 2֯ sensory neurons, associated interneurons, 1 sensory neurons from Neural Crest Floorplate: Ventral signal→ Sonic Hedgehog Motor neurons, ventral roots of spinal cord, associated interneurons SHH is also responsible for stimulating the formation of the MHP (median hinge point) 32 16  19/01/2024 Rostral-caudal axis Signals from the anterior visceral endoderm, the prechordal plate, and the notochord: - Otx-2 in the future forebrain and midbrain regions - Gbx-2 in the hindbrain and spinal cord Later in development: 1. Rostral neural tube forms 3 primary brain vesicles - prosencephalon (forebrain) - mesencephalon (midbrain) - rhombencephalon (hindbrain) 2. Caudal→ spinal cord 33 Cell Lineages of the Central Nervous System Neuroepithelial stem cells Neuroepithelial stem cells Radial precursor cells Radial precursor cells Neurogenesis (after patterning neural tube) Radial glial cells - One daughter cell remains as radial precursor cell - The other, radial glial cell neuroblast and ultimately a neuron IPC two neuroblasts Neuron-glial cell switch: downregulation of neuregulin, secretion of neurogenesis-inhibitory cytokines, change in the growth factor environment, activation of progliogenic transcription factors… Gliogenesis - Radial precursor cell OPC Oligodendrocytes APC Astrocytes (2 lines) 34 17  19/01/2024 Neuronal migration in the CNS Neuroblast migrate toward the periphery by following set patterns Radial glial cells→ postmitotic neurons wrap themselves around the radial glial cells and use them as guides on their migrations - Inside–out pattern→ cortex - Out-inside pattern→ cerebellum 35 Instructions Join at vevox.app Or search Vevox in the app store ID: 128-025-112 Where opportunity creates success 36 18  19/01/2024 Join: vevox.app ID: 128-025-112 POLL OPEN Which of these cells do neural crest cells not differentiate into? 1. Adrenal gland cell precursors 2. Dorsal root ganglia 3. Microglia 4. Pigment cells 5. Sympathetic ganglia 37 Join: vevox.app ID: 128-025-112 POLL OPEN Which genetic signal, from the floorplate (ventral) compartment, stimulates the differentiation into motor neurons and ventral roots of the spinal cord? 1. Bone morphogenetic protein (BMP) 2. Gbx2 3. Hox 4. Otx2 5. Sonic hedgehog (SHH) 38 19  19/01/2024 10 minute break 39 2 minute neuroscience 40 20  19/01/2024 Early brain development 41 Regions of the brain Brain starts to form distinct regions: first three, then 5. Cranial flexures impact on brain development: allowing expansion of the cavities and creating regions and boundaries. – Pontine flexure separates the walls and forms the floor of the 4th ventricle – Cephalic flexure creates the bend between the prosencephalon and the rest of the CNS 42 21  19/01/2024 Differentiation of the Forebrain Retina part of forebrain, not PNS Telencephalon: cerebral hemispheres, olfactory bulbs, basal telencephalon Diencephalon: thalamus and hypothalamus Axons extend from developing forebrain to other parts of nervous system – Cortical white matter – Corpus callosum – Internal capsule Forebrain structure-function relationships – Cerebral cortex Analyze sensory input and command motor output – Thalamus: gateway to the cortex 43 Differentiation of the Midbrain Midbrain structure-function relationships – Contains axons descending from cortex to brain stem and spinal cord Example: corticospinal tract – Information conduit from spinal cord to forebrain and vice versa, sensory systems, control of movements – Tectum—superior colliculus (receives sensory info from eye), inferior colliculus (receives sensory info from ear) – Tegmentum Substantia nigra (‘black substance’ – dopamine-producing neurons) and red nucleus—control voluntary movement 44 22  19/01/2024 Differentiation of the Rostral and Caudal Hindbrain Hindbrain structure-function relationships – Cerebellum: movement control – Pons: switchboard connecting cerebral cortex to cerebellum – Cochlear nuclei: project axons to different structures (e.g., inferior colliculus) – Decussation: crossing of axons from one side to the other 45 Differentiation of the spinal cord Dorsal horn→ sensory inputs Ventral horn→ motor outputs 6 tracts – Most are one way – Bring information from or to brain 46 23  19/01/2024 Putting the pieces together 47 Mapping fetal brain development 48 24  19/01/2024 How are boundaries set up? Neural tube is organized in repeating units called neuromeres. Different gene expression patterns set up cell identity: Gbx2 is one of the first brain signals Otx2 sets up forebrain identity Hox genes (homeotic selector genes): crucial genes in development for setting up expression domains. They control expression of other genes which will then further specify identity and patterning Hox gene expression controlled by different cis regulatory molecules 49 Construction of neural circuits 50 25  19/01/2024 Neuronal polarization Neurons→ polarized epithelial cells – Basolateral domain secrete proteins (intercellular communication) – Apical domain absorb molecules (cilia or microvilli) Neuronal polarization→ distinguish its domains (after neurogenesis) 1. Neurites (small processes of developing neurons) 2. Axons → secretion 3. Dendrites → signal transduction PAR family as polarity regulators 51 Axon growth Growth cone→ specialized structure at the tip of the axon – Determine direction of the axon – Guide the extension of the axon in that direction Lamellipodium and filipodia Cues in their target to guide the process Transform in the presynaptic ending for an axon or the terminal domain of a dendrite (extend long distances) Growth cones changes shape when reach the target 52 26  19/01/2024 Molecular mechanism of growth cone motility Controlled rearrangement of the cytoskeleton – Actin→ changes in shape – Microtubule→ elongation of the axon ATP-dependent 53 Actin treadmilling 375 amino acid polypeptide carrying a associated molecule of ATP Actin subunits assemble head-to tail to create flexible polar filaments The polymer maintains constant length and can move across the cytosol Two ends of an actin filament→ Plus end (faster dynamics) and minus end (slower dynamics) Actin-ATP associates to plus end ADP-actin dissociate from minus end 54 27  19/01/2024 Actin binding proteins Regulate polymerisation and de- polymerisation Mediate assembly and membrane anchoring of actin filaments Found in the inner surface of the growth cone plasma membrane 55 Growth cones 56 28  19/01/2024 Instructions Join at vevox.app Or search Vevox in the app store ID: 128-025-112 Where opportunity creates success 57 Join: vevox.app ID: 128-025-112 POLL OPEN What part of a developing neuron guides extension of the axon to the target region? 1. Cytosol 20.83% 2. Endoplasmic reticulum 22.92% 3. Growth cone 22.92% 4. Nucleolus 14.58% 5. Soma 18.75% Sample Results 58 29  19/01/2024 Join: vevox.app ID: 128-025-112 POLL OPEN Which early brain region does the corticospinal tract develop from? 1. Cerebellum 14% 2. Diencephalon 32% 3. Mesencephalon 16% 4. Rhombencephalon 20% 5. Telencephalon 18% Sample Results 59 2 minute break 60 30  19/01/2024 Microtubule assembly Consist of subunits called tubulin heterodimers (α- and β-tubulin) Dimers build up Protofilament polarised Hollow tubes build from 13 protofilaments Lateral contacts between monomers of the same type (straight and stiff structures) Disassembly and assembly occur simultaneously→ Dynamic instability – Catastrophe→ Hydrolysis more rapidly than subunit addition (cap lost/ begin to shrink) – Rescue→ while shrinking enough GTP-containing subunits to form a new cap – Dependent on GTP hydrolysis (on β-tubulin) 61 Microtubule associated proteins MAPs with one domain that binds to the microtubule surface and another that projects outward Found in the axon shaft Dynein and kinesin→ moving molecules and organelles up and down neuronal processes 62 31  19/01/2024 Signal for axon guidance Presence of specific “cues” (proteins) that cause the growth cone to move in a particular direction Initiate intracellular signalling→ alter the cytoskeleton 1. EM molecules and integrins receptors 2. Cell adhesion molecules 3. Cadherins 4. Ephrins 63 EM molecules and integrins receptors Laminins, collagens and fibronectins – Can be secreted by the cell itself or by its neighbours – Role in the embryonic PNS→ fill the spaces between mesenchymal cells and organised the basal lamina – Role in the embryonic CNS→ are found in the extracellular space Integrins receptors – Cell surface receptors – Binds to these molecules 64 32  19/01/2024 CAMs and Cadherins Found on growth cones as well as on surrounding cells and target Dual function→ ligands and receptors Cadherins→ determinate the final target selection Signal transduction – CAMs→ cytoplasmatic kinases pathway – Cadherins→ β-catenin pathway 65 Ephrins Tyrosine kinase receptors (Eph) Role in synaptogenesis and the growing of the immature axons Depending on the nature of signal transduction – Growth-promoting – Growth-limiting 66 33  19/01/2024 Chemoattraction and chemorepulsion How do the axons choose the appropriate target? 1. Chemoattractants→ netrins High homology with EMC (laminins) Receptors→ DCC, Unc5 2. Chemorepellents→ CNS myelin and semaphorins Bound to the cell surface or ECM Receptors→ plexin, neuropilin 67 Commissural Neuron Migration Commissural Neurons migrate across the midline Netrins – Chemoattractive signal in the spinal cord ventral midline – Guide the growth of spinothalamic axons – Localized in the floorplate of neural tube Slit and Robo prevent the axon returning back 68 34  19/01/2024 Dendritic growth Maintenance, guided growth and local branching of dendrites Chemoattractive and chemorepellant signalling Sema3A – Repels the axons of developing neurons – Attract for the dendrites of the same cells (via neuropilin recptors) Notch and BDNF act as a positive signal for cortical dendritic branching 69 Dendritic tiling Modulation of dendritic growth where each dendritic arbor occupies appropriate space – Avoiding dendrites from the same neuron – Adequate coverage for a particular region DSCAM genes→ dual repulsion 1. Responding to molecular signals from its own dendrites 2. Responding from the dendrites of other neurons 70 35  19/01/2024 Synapse formation Restrictions in synapse formation – CNS neurons don’t synapse with glia – PNS neurons don’t synapse with connective cells Relative promiscuity Overlapping subset of molecules that regulate general aspect in synapse formation Secreted signal implicated in this process→ growth factors and neurotransmitters 71 Molecular mechanism 1. Local recognition pre and post synaptic – Cadherin and protocadherin – Accumulation of synaptic vesicles 2. Adhesive signalling – SynCAM, Neurexin, Neuroligin, Neuregulin – Differentiation of active zone and postsynaptic density – Clustering postsynaptic receptors 3. Localizing molecules – Synaptic vesicles (neurexin) – Voltage-gated Ca²⁺ channels (neurexin) – Neurotransmitters receptors (neuroligin) 72 36  19/01/2024 Regulation of neuronal connections Trophic interaction→ dependency between neurons and their target Neurotrophic factors→ regulate differentiation, growth and survival in nearby cells Production of initial surplus of nerve cells Survival depends on location, species, and activity → large numbers of neurons die because they do not make synapses (apoptosis) 73 Modulation of neuronal connections 1. Convergence→ number of inputs to a target cell 2. Divergence→ number of connection made by a neuron Polyneuronal innervation (embryonic stage) Synapse elimination→ reduction in number of axonal inputs to the target cell Synapse contacts increase in PNS and CNS along the life 74 37  19/01/2024 Neurotrophins and Disease Neurotrophic factors have been liked with postnatal disease. Huntington’s Disease Parkinson’s Decreased Brain Derived Neurotrophic Factor Decreased Glial Derived (BDNF) Neurotrophic Factor (GDNF) and BDNF? Death of striatal neurons GDNF promotes survival of Involuntary muscle movements and cognitive dopaminergic neurons problems BDNF required for neural response to dopamine 75 38